Temporal trends (1972–2017) and spatial differences of persistent halogenated aromatic hydrocarbons in osprey eggs in Finland

Time trends and regional differences of polychlorinated dibenzo-p-dioxins and -furans (PCDD/Fs), polychlorinated biphenyls (PCBs), polychlorinated naphthalenes (PCNs), DDTs, polybrominated biphenyls (PBBs) and polybrominated diphenylethers (BDEs) were studied in unhatched osprey eggs collected by bird ringers in 1972–2017 from four areas in Finland. Two study areas were from Baltic Sea, Northern Quark and Finnish Archipelago Sea, while the two others were inland lake areas, eutrophicated Lake Vanajanselkä affected by industrial emissions, and Pristine SW Lake Area. The highest concentrations of most compound groups were in Lake Vanajanselkä consistent with high emissions, the predominance of bream as a prey, and higher concentrations in bream compared to other prey fish. Concentrations of all chlorinated compounds decreased significantly in all study areas. Average annual decreases were ∑PCDD/F 2.3–4.9%, ∑PCB 2.2–4.2%, ∑PCN 2.6–7.0% and ∑DDT 7.1–9.5%, primarily in line with decreased levels in prey fish. From 1972 PBBs and BDEs increased significantly until 1990s declining rapidly thereafter. PCDD/F congener profile was dominated by 2,3,4,7,8-PeCDF, except in Lake Vanajanselkä by 1,2,3,6,7,8-HxCDD. PCB congener profile was dominated by PCB 153 in all study areas, followed by PCB 180 and PCB 138. Among dioxin-like compounds PCBs contributed 82%, PCDDs 14% and PCDFs 4% to toxic equivalent quantity (∑TEQ). PCB 126 contributed most to ∑TEQ, followed by 1,2,3,7,8-PeCDD. BDE 47 being the dominant BDE congener, followed by BDE 100. ∑DDT concentrations were relatively similar across all study areas, with DDE contributing about 90%. Productivity of chicks per active nest was significantly decreased in Lake Vanajanselkä, and the likely explanation is embryotoxicity of dioxin-like compounds. It is plausible that dioxin-like compounds influenced embryonic survival among highly exposed ospreys prior to 2010, especially in Lake Vanajanselkä and Northern Quark. However, decreased survival due to DDE-induced eggshell thinning seems unlikely after 1985, and BDE levels were below those potentially causing adverse effects.


Chemicals and standards
Solvents and solid reagents used were of the highest commercial quality available.Both 12 C and 13 C standard compounds for POPs were obtained from Wellington or Cambridge Isotope Laboratories.However, the number of 13 C-labelled internal standards slightly increased over the years for PCBs (2004: 17; 2018: 23) and BDEs (2004: 6;2018: 8).

Extraction and clean-up of POPs
Egg samples (2-10 g depending on the availability of sample) were spiked with 13 C-labelled internal standards, mixed with concentrated hydrochloric acid (15 ml) and heated to 100℃.After cooling, ultrapurified water (30 ml) was added to the sample and extracted in a separatory funnel with diethyl ether (30 ml).n-hexane (30 ml) was added and extraction continued.Organic phase was separated.The extraction of aqueous phase with diethyl ether and n-hexane was repeated twice more.The combined organic phase was washed with ultra-purified water.The water fraction was then rejected.The organic phase was dried with sodium sulfate and filtrated through a cotton plug.The solvent was evaporated almost to dryness and reconstituted to pure n-hexane before column clean-up.Samples were defatted with multilayer silica column cleanup and fractionated by activated carbon column as previously described [1].PCBs, PBDEs, PBBs and DDTs were collected from carbon column in the forward eluted fraction 1 (F1) while PCDD/Fs, non-ortho-PCBs and PCNs were collected in backwards eluted fraction 2 (F2).F1 was concentrated to 500 µl of hexane and F2 to 15 µl of nonane before gas chromatographyhigh resolution mass spectrometry (GC-HRMS) or gas chromatography triple quadrupole mass spectrometer (GC-MS/MS) analysis.

Instrumental analysis of POPs
In 2004 and 2007 POPs were analyzed with Waters Autospec Ultima GC-HRMS and in 2018 with Agilent 7010 GC-MS/MS.GC column was DB-5MS UI (J&W Scientific, 60m, ID 0.25 mm, 0.25 μm film) in both cases.All groups of POPs were analyzed in separate GC-HRMS or GC-MS/MS run programs except for PCDD/Fs and non-ortho-PCBs that were analyzed in the same run.The status of both GC-HRMS and GC-MS/MS was assessed daily, and the instruments were calibrated and serviced regularly.

Quality control in POP analysis
Two blank samples were included in the batch of samples.Laboratory of chemistry at the Finnish Institute for Health and Welfare is an accredited testing laboratory T077 by Finnish Accreditation Services (FINAS) since 1996.Scope of accreditation includes PCDD/Fs, PCBs and PBDE from egg samples.Finnish Institute for Health and Welfare is also the National Reference Laboratory (NRL) for Halogenated POPs in Feed and Food in Finland.

Toxic equivalency factors and nomenclature of congeners
Toxic equivalency factors (TEFs) [2] used for calculation of toxic equivalency quantities (TEQs) of dioxinlike compounds are shown in

Table B . Groups of POPs and individual congeners analyzed. POPs
with detection rates < 20% were excluded from statistical analysis with the exception of PCDD/Fs for which all congeners were included.Congeners marked with '*' had 13 C-labelled internal standard.

Table C . WHO 2005 toxic equivalency factors (TEFs) of dioxin-like congeners
Table C and IUPAC names of numbered PCB, PBB, BDE and PCN congeners analyzed in this study in Tables D-G.